Proteases from purulent sputum. Purification and properties of the elastase and chymotrypsin-like enzymes.

Twumasi, D Y; Liener, Irvin E.

doi:10.1016/s0021-9258(18)71845-0

Cited by 110 publications

(8 citation statements)

References 43 publications

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“…Such bands have been demonstrated to represent distinct, catalytically active HLE isoforms, which differ in their carbohydrate content and have been resolved by SDS-PAGE. − Moreover, the self-cleavage of elastase from murine and human neutrophils generates variants whose catalytic activity was shown by means of a biotinylated probe . The occurrence of the pattern of three elastase bands (Figure ) has been observed in several previous studies. − ,− Thus, it can be attributed to either differences in the carbohydrate contents or to an autoprocessing of elastase. Compared to the Coomassie Brilliant Blue staining, probe 6 produced a much stronger labeling of HLE (Figure A versus B).…”

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confidence: 73%

A BODIPY-Tagged Phosphono Peptide as Activity-Based Probe for Human Leukocyte Elastase

Schulz-Fincke

Blaut

Braune

et al. 2018

ACS Med. Chem. Lett.

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Human leukocyte elastase plays a crucial role in a variety of inflammatory disorders and represents an important subject of biomedical studies. The chemotype of peptidic phosphonates was employed for the design of a new activity-based probe for human leukocyte elastase. Its structure combines the phosphonate warhead with an adequate peptide portion and a BODIPY fluorophore with a clickable ethinylphenyl moiety at position. The probe was assembled by copper-catalyzed alkyne-azide 1,3-dipolar cycloaddition. It was characterized as an active site-directed elastase inhibitor exhibiting a second-order rate constant of inactivation of 88400 Ms. The suitability of as a fluorescent probe for human leukocyte elastase was demonstrated by in-gel fluorescence analysis. Labeling experiments and inhibition data toward a panel of related proteases underlined the selectivity of the probe for the targeted leukocyte elastase.

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confidence: 73%

A BODIPY-Tagged Phosphono Peptide as Activity-Based Probe for Human Leukocyte Elastase

Schulz-Fincke

Blaut

Braune

et al. 2018

ACS Med. Chem. Lett.

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“…HLE is known to contain a number of isozymes that are thought to differ only in carbohydrate content (Twumasi & Liener, 1977;Baugh & Travis, 1976). The carbohydrate content reported has ranged from 9-12% (Twumasi & Liener, 1977) to as high as 21% (Baugh & Travis, 1976). The most predominant isozymes present in HLE are isozymes 3 and 4 (Green et al, 1991).…”

mentioning

confidence: 99%

Electrospray ionization mass spectrometry as a mechanistic tool: Mass of human leukocyte elastase and a .beta.-lactam-derived E-I complex

Knight¹,

Swiderek²,

Sakuma³

et al. 1993

Biochemistry

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We have utilized liquid chromatography electrospray ionization mass spectrometry (ESI-MS) to probe the nature of the covalent E-I complex of human leucocyte elastase (HLE) and a beta-lactam. The mass spectrum of HLE isozyme 4 displayed one major and two minor components with masses of 25,202, 25,043, and 24,522 Da, respectively. Isozyme 3 displayed three components, with masses of 25,180, 24,030, and 24,523 Da. These data suggest that the isozymes differ in the type and not the content of carbohydrate. The minor components represent decreases in carbohydrate content. Inactivation of isozyme 4 with trans-4-(ethoxycarbonyl)-3-ethyl-1-[(4-nitrophenyl)sulfonyl]-azetidin -3-one increased the mass of the three components by that of the parent compound. Similar results were obtained with the mixture of HLE isozymes. These observations demonstrate that HLE does not catalyze the beta-elimination of p-nitrophenylsulfinate as Firestone et al. [(1990) Tetrahedron 46, 2255) suggested. In addition, it suggests that a "double hit" of both the active-site serine and histidine is not required to form a stable acyl-enzyme. Noncovalent complexes between HLE and either the tight-binding secretory leucoprotease inhibitor (SLPI) or a slow tight-binding peptide difluoroketone inhibitor were not observed by ESI-MS. SLPI displayed a mass of 11,710 Da in the absence and presence of HLE. These data demonstrate the utility of ESI-MS to probe the mechanism of inhibition of enzymes by mechanism-based inhibitors.

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“…It is known that several HNE isoforms can be resolved by SDS–PAGE and that these catalytically active forms differ only in their carbohydrate content. Moreover, the self-cleavage of elastase from murine and human neutrophils was shown to generate variants with different catalytic activities . Because the elastase used in our study was prepared from human neutrophils, we assume that the three bands correspond to HNE isoforms with different glycosylation patterns or are caused by autocatalytic cleavage.…”

Section: Resultsmentioning

confidence: 99%

Design of an Activity-Based Probe for Human Neutrophil Elastase: Implementation of the Lossen Rearrangement To Induce Förster Resonance Energy Transfers

et al. 2018

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Human neutrophil elastase is an important regulator of the immune response and plays a role in host defense mechanisms and further physiological processes. The uncontrolled activity of this serine protease may cause severe tissue alterations and impair inflammatory states. The design of an activity-based probe for human neutrophil elastase reported herein relies on a sulfonyloxyphthalimide moiety as a new type of warhead that is linker-connected to a coumarin fluorophore. The inhibitory potency of the activity-based probe was assessed against several serine and cysteine proteases, and the selectivity for human neutrophil elastase (K = 6.85 nM) was determined. The adequate fluorescent tag of the probe allowed for the in-gel fluorescence detection of human neutrophil elastase in the low nanomolar range. The coumarin moiety and the anthranilic acid function of the probe, produced in the course of a Lossen rearrangement, were part of two different Förster resonance energy transfers.

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Proteases from purulent sputum. Purification and properties of the elastase and chymotrypsin-like enzymes.

Cited by 110 publications

References 43 publications

A BODIPY-Tagged Phosphono Peptide as Activity-Based Probe for Human Leukocyte Elastase

A BODIPY-Tagged Phosphono Peptide as Activity-Based Probe for Human Leukocyte Elastase

Electrospray ionization mass spectrometry as a mechanistic tool: Mass of human leukocyte elastase and a .beta.-lactam-derived E-I complex

Design of an Activity-Based Probe for Human Neutrophil Elastase: Implementation of the Lossen Rearrangement To Induce Förster Resonance Energy Transfers

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